Hydrogen peroxide can be used for fumigating isolators or entire rooms in order to destroy microorganisms which are possibly to be found therein. The hydrogen peroxide in gas form condenses on the fumigated surfaces, as a between 30% and 35% saturated solution. Prior to the start of manufacturing procedures, quality control investigations in respect of sterility or other operations in the fumigated clean room areas, they are sterily ventilated, whereafter levels of concentration of between 0.3 and 6 ppm, in general below 1 ppm, of germs remain in the air. Investigations in regard to such air-borne germs which are possibly still present is effected with air-borne germ collecting devices operating on the basis of the impaction or rotation principle, with the germ-bearing particles being deposited on agar surfaces.
Surprisingly the small amounts of hydrogen peroxide vapors are concentrated in the course of collecting 1000 liters of air in casein soya peptone agar, in accordance with United States Pharmakopoeae, 8th Supplement, USP-NF, <1116>, 4426-4431, on concentrations of over 100 ppm in agar. Spores are already restrained by levels of hydrogen peroxide concentration of 10 ppm and vegetative cells and microorganisms are already restrained by an even more markedly lower concentration of hydrogen peroxide. That adversely affects to a considerable degree the detectability of microorganisms which are still present.
Normal agar media have a contamination rate of about 0.1%, that is to say 1 non-sterile unit among 1000, and therefore do not comply with the notion of sterility which allows only one non-sterile unit among 106 (one million). As the media used for the investigatory procedure are employed to investigate clean rooms which were previously rendered germ-free by fumigation, it would be desirable for no germs to be introduced with the agar materials for testing for freedom from germs, and therefore the expectation is for sterile media.
For conducting detection procedures of the above-indicated kind, it is possible to use media such as the Baird-Parker medium (Journ. Applied Bacteriology 25:12, 1962, Baird-Parker) with 1% of sodium pyruvate, being a sodium salt of pyruvic acid or 2-oxopropionic acid, which inter alia are suitable for the isolation of Staphylococcus aureus after heat damage or storage of frozen or dried vegetative cells. The disadvantage of Baird-Parker agar is the low level of stability and durability of the finished agar medium.
The addition of catalase to this and other media for improving the growth of bacteria from the air is also known, in which respect reference may be made to Journ. Applied and Environmental Microbiology 57: 2775-2776, 1991, Balkumar Marthi. Catalase breaks down hydrogen peroxide into water and oxygen. Catalase however is inactivated at 55° C., which presupposes drawing off agar at markedly below 55° C. That however is scarcely a viable option because of gelling of the agar at temperatures around 50° C. In addition the oxygen resulting from the hydrogen peroxide causes bubbles and cracks in the agar, which causes extreme difficulty in detecting colonies which grow on the agar.
Also known is D/E-agar which neutralises a wide range of antiseptic and disinfecting chemicals including quaternary ammonium compounds, phenol, iodine and chlorine compounds, mercury (Merthiolate), formaldehyde and glutaraldehyde (Difco Handbook, D/E-Agar). That Difco Handbook does not describe neutralisation of hydrogen peroxide. The disadvantage of D/E-agar is the short durability life of the ready agar medium of only about two and a half months and the changes in the medium in the event of radiation doses of 16-25 kgray, which are necessary for reliable gamma-sterilisation.
In addition D/E-agar is not very stable in respect of pH and at a pH of 7.6±0.2, is already of a very high pH-value. Upon drifting towards even higher pH-values such as 8.0, namely only 0.2 above the upper limit of the range specified above, the situation already involves marked restraints in respect of germs to be detected.